Biomedical subjects

Blake C Ellis

Publications and source records attributed to Blake C Ellis.

4 recordsLinked to original sources

PubMed2006

A dolphin peripheral blood leukocyte cDNA microarray for studies of immune function and stress reactions.

A microarray focused on stress response and immune function genes of the bottlenosed dolphin has been developed. Random expressed sequence tags (ESTs) were isolated and sequenced from two dolphin peripheral blood leukocyte (PBL) cDNA libraries biased towards T- and B-cell gene expression by stimulation with IL-2 and LPS, respectively. A total of 2784 clones were sequenced and contig analysis yielded 1343 unigenes (archived and annotated at ). In addition, 52 dolphin genes known to be important in innate and adaptive immune function and stress responses of terrestrial mammals were specifically targeted, cloned and added to the unigene collection. The set of dolphin sequences printed on a cDNA microarray comprised the 1343 unigenes, the 52 targeted genes and 2305 randomly selected (but unsequenced) EST clones. This set was printed in duplicate spots, side by side, and in two replicates per slide, such that the total number of features per microarray slide was 19,200, including controls. The dolphin arrays were validated and transcriptomic profiles were generated using PBL from a wild dolphin, a captive dolphin and dolphin skin cells. The results demonstrate that the array is a reproducible and informative tool for assessing differential gene expression in dolphin PBL and in other tissues.

Animals↗

PubMed2005

Establishment of epidermal cell lines derived from the skin of the Atlantic bottlenose dolphin (Tursiops truncatus).

The Atlantic bottlenose dolphin (Tursiops truncatus), a marine mammal found off the Atlantic coast, has become the focus of considerable attention because of an increasing number of mortality events witnessed in this species over the last several years along the southeastern United States. Assessment of the impact of environmental stressors on bottlenose dolphins (BND) has been difficult because of the protected status of these marine mammals. The studies presented herein focused on establishing epidermal cell cultures and cell lines as tools for the in vitro evaluation of environmental stressors on BND skin. Epidermal cell cultures were established from skin samples obtained from Atlantic BND and subjected to karyotype analysis. These cultures were further characterized using immunohistochemical methods demonstrating expression of cytokeratins. By two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), we observed that the proteomic profile of BND skin tissue samples shared distinct similarities with that of skin-derived cultures. Epidermal cell cultures were transfected with a plasmid encoding the SV40 small t- and large T-antigens, as well as the neomycin-resistance gene. Five neomycin-resistant clones were isolated and expanded, and all of them proliferated at a faster rate than nontransfected BND epidermal cultures, which exhibited signs of senescence. Cell lysates prepared from two transfected clones were shown to express, by Western blot analysis, both SV40 tumor antigens. These experimental results are consistent with the concept that transfected clones expressing SV40 tumor antigens represent immortalized BND cell lines. Epidermal cell lines derived from Tursiops truncatus will provide a unique tool for studying key features of the interaction occurring between dolphins and the environment in which they live at their most crucial interface: the skin.

Animals↗

PubMed2004

Role of acyl-coenzyme a: cholesterol acyltransferase activity in the processing of the amyloid precursor protein.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by progressive memory deficit, cognitive impairment, and personality changes accompanied by specific structural abnormalities in the brain. Deposition of amyloid-beta (Abeta) peptide into senile plaques is a consistent feature of the brains of patients affected by AD. Studies with both animal and cellular models of AD have shown that cholesterol homeostasis and distribution regulate Abeta generation. We have provided genetic, biochemical, and metabolic evidence that implicates intracellular cholesterol distribution, rather than total cholesterol levels, in the regulation of Abeta generation. This minireview focuses on the role of acyl-coenzyme A: cholesterol acyltransferase activity (ACAT) in Abeta generation. In genetically mutant cell lines that overproduce cholesterol but cannot synthesize cholesteryl esters (CEs) because of deficient ACAT activity, Abeta production is almost completely inhibited. Acyl-coenzyme A: cholesterol acyltransferase activity (ACAT) inhibitors, currently being developed for the treatment and prevention of atherosclerosis, reduce CE levels and Abeta generation by up to 50% in cell culture models of AD. Future mechanistic and transgenic animal studies are needed to evaluate the potential use of ACAT inhibitors in the therapeutic treatment or prevention of AD.

Acyl Coenzyme A↗

PubMed2003

Ceramide stabilizes beta-site amyloid precursor protein-cleaving enzyme 1 and promotes amyloid beta-peptide biogenesis.

The lipid second messenger ceramide regulates several biochemical events that occur during aging. In addition, its level is highly elevated in the amyloid-burdened brains of Alzheimer's disease patients. Here, we analyzed the impact of aberrant ceramide levels on amyloid beta-peptide (Abeta) generation by using a cell-permeable analog of ceramide, C6-ceramide, and several biochemical inhibitors of the sphingomyelin/glycosphingolipid biosynthetic pathway. We found that C6-ceramide increased the biogenesis of Abeta by affecting beta-but not gamma-cleavage of the amyloid precursor protein. Similarly to C6-ceramide, increased levels of endogenous ceramide induced by neutral sphingomyelinase treatment also promoted the biogenesis of Abeta. Conversely, fumonisin B1, which inhibits the biosynthesis of endogenous ceramide, reduced Abeta production. Exogenous C6-ceramide restored both intracellular ceramide levels and Abeta generation in fumonisin B1-treated cells. These events were specific for amyloid precursor protein and were not associated with apoptotic cell death. Pulse-chase and time-course degradation experiments showed that ceramide post-translationally stabilizes the beta-secretase BACE1. Taken together, these data indicate that the lipid second messenger ceramide, which is elevated in the brains of Alzheimer's disease patients, increases the half-life of BACE1 and thereby promotes Abeta biogenesis.

Amyloid Precursor Protein Secretases↗